Abstract
Expression of all Yersinia pathogenicity factors encoded on the virulence plasmid, including the yop effector and the ysc type III secretion genes, is controlled by the transcriptional activator LcrF in response to temperature. Here, we show that a protein- and RNA-dependent hierarchy of thermosensors induce LcrF synthesis at body temperature. Thermally regulated transcription of lcrF is modest and mediated by the thermo-sensitive modulator YmoA, which represses transcription from a single promoter located far upstream of the yscW-lcrF operon at moderate temperatures. The transcriptional response is complemented by a second layer of temperature-control induced by a unique cis-acting RNA element located within the intergenic region of the yscW-lcrF transcript. Structure probing demonstrated that this region forms a secondary structure composed of two stemloops at 25°C. The second hairpin sequesters the lcrF ribosomal binding site by a stretch of four uracils. Opening of this structure was favored at 37°C and permitted ribosome binding at host body temperature. Our study further provides experimental evidence for the biological relevance of an RNA thermometer in an animal model. Following oral infections in mice, we found that two different Y. pseudotuberculosis patient isolates expressing a stabilized thermometer variant were strongly reduced in their ability to disseminate into the Peyer's patches, liver and spleen and have fully lost their lethality. Intriguingly, Yersinia strains with a destabilized version of the thermosensor were attenuated or exhibited a similar, but not a higher mortality. This illustrates that the RNA thermometer is the decisive control element providing just the appropriate amounts of LcrF protein for optimal infection efficiency.
Highlights
Pathogenic yersiniae, including Y. pestis, the causative agent of the bubonic plague, and the two enteric species Y. enterocolitica and Y. pseudotuberculosis which cause gut-associated diseases such as enteritis, diarrhea and mesenterial lymphadenitis express different sets of virulence factors important for different stages of the infection process [1,2]
One important set of thermo-induced virulence factors is encoded on the 70 kb Yersinia virulence plasmid pYV [10]
Many important virulence genes remain silent at moderate temperatures in external environments and are rapidly and strongly induced by a sudden temperature upshift sensed upon host entry
Summary
Pathogenic yersiniae, including Y. pestis, the causative agent of the bubonic plague, and the two enteric species Y. enterocolitica and Y. pseudotuberculosis which cause gut-associated diseases (yersiniosis) such as enteritis, diarrhea and mesenterial lymphadenitis express different sets of virulence factors important for different stages of the infection process [1,2]. Some of the early stage virulence factors, including the primary internalization factor invasin of both enteric Yersinia species, are mostly produced at moderate temperatures to allow efficient trespassing of the intestinal epithelial barrier shortly after infection [4,5,6]. These virulence genes are controlled by RovA, an intrinsic protein thermometer, which undergoes a conformation change upon a temperature shift from 25uC to 37uC, that reduces its DNA-binding capacity and renders it more susceptible to proteolysis [7,8,9].
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